PROJECT 1 SUMMARY Existing antibody-based interventions for cancer rely on passive delivery of the circulating drug that depends on a large concentration gradient across the semi-permeable wall of blood vessels to get inside tumors. The forces driving the drug from the bloodstream into tumors result in sub-optimal delivery and poor access to tumor cells, in part because endothelial cells (EC) forming the vascular wall constitute a significant, limiting barrier. We intend to boost precision delivery into primary and metastatic tumors by overcoming the vascular EC barrier through a highly precise, active transport pathway that we discovered through proteomic imaging and named the caveolae pumping system. The approach proposed here to enhance delivery goes well beyond typical so- called `active targeting' of antibodies. Our lead humanized antibody against Annexin A1 (hAnnA1), a tumor- specific antibody concentrated in EC caveolae, not only binds its intended target but actually is the first antibody to penetrate solid tumors actively, rapidly and specifically. It does so even at very low dosages and reaches unprecedented intra-tumoral concentrations well beyond the highest blood levels after injection. It is now time to test this unprecedented immunotargeting in humans. We propose here to develop a radiolabeled humanized mAnnA1 to detect and destroy primary and metastatic lesions. Major aims of this project are to: 1-2) evaluate in vivo delivery and efficacy of novel caveolae-targeting radioimmunoconjugates in hard-to-treat metastatic tumor models and canine cancer patients; 3) test the effectiveness of caveolae pumping in human tumor blood vessels using PDX and novel human IVM tumor models; and 4) translate hAnnA1 towards clinical testing. We will determine here the degree to which caveolae can be targeted to pump radiolabeled antibodies across vascular EC to concentrate them inside solid tumors as a means to improve image-guided drug delivery and enhance their efficacy. Antibodies will be provided by Core B and radiolabeled with assistance provided by Core D for preclinical work. Several mammary tumor models will be used to assess the ability of radiolabeled hAnnA1 to target primary and metastatic tumors via imaging services provided by Core C. We will study intravenously injected hAnnA1 with advanced multimodality in vivo imaging to quantify and optimize precision transvascular delivery and tumor penetration. As the utility of caveolae targeting in humans requires AnnA1 expression in tumor EC caveolae, we will use Core C services to compare vascular expression of AnnA1 in preclinical tumor models and human solid tumors. To translate our findings for clinical testing, Core B will oversee cGMP production of hAnnA1. Following radiolabeling in Core D, Project 3 will conduct a first-in-human imaging trial as a direct result of successful translation of our findings from Project 1 into the clinic.